BIOLOGY OF REPRODUCTION 63, 526–531 (2000)
Determination of Seasonality in Southern Hairy-Nosed Wombats
(Lasiorhinus latifrons) by Analysis of Fecal Androgens1
Ross A. Hamilton,3,4 Peter G. Stanton,3 Liza O’Donnell,3 Vernon R. Steele,4 David A. Taggart,4
and Peter D. Temple-Smith2,5
Prince Henry’s Institute of Medical Research,3 Clayton, Victoria 3168, Australia
Department of Anatomy,4 Monash University, Clayton, Victoria 3168, Australia
Conservation & Research Unit,5 Zoological Parks & Gardens Board, Parkville, Victoria 3052, Australia
ABSTRACT
INTRODUCTION
Little is known about the reproductive biology of Australia’s
critically endangered northern hairy-nosed wombat (Lasiorhinus
krefftii), largely due to its cryptic nature and the difficulty in
accessing the small remaining population of about 70 animals.
Using the noninvasive technique of fecal steroid analysis, we
have examined the endocrinology of the more common yet
closely related southern hairy-nosed wombat (Lasiorhinus latifrons). The aims of this study were to 1) develop and validate
fecal androgen analysis in this species, 2) examine and compare
seasonal differences in fecal and plasma androgens in male
wombats, and 3) correlate seasonal differences in androgens
with changes in male accessory glands (prostate and bulbourethral gland). Fecal androgens were extracted in ether; concentrated; separated by HPLC into testosterone (T), dihydrotestosterone (DHT), and 5a-androstane-3a,17b-diol (Adiol) fractions;
and quantitated by RIA. The concentrations of androgens in fecal
pellets from 14 wild southern hairy-nosed wombats as determined by RIA varied over the range 6.6–25.0 ng/g dry weight
for T, 4.0–24.2 ng/g dry weight for DHT, and 0–34.8 ng/g dry
weight for Adiol. For each androgen, a highly significant linear
correlation was observed between plasma and fecal concentrations. When individuals were grouped into either breeding season (pellets collected between August–November) or nonbreeding season (collected between February–April), significant ( P ,
0.05) differences between seasons were observed for both plasma and fecal T, plasma DHT, and fecal Adiol. For all androgens,
the mean fecal and plasma concentrations were higher during
the breeding season than the nonbreeding season. A significant
(P , 0.001) correlation was observed between fecal T and prostate weight, while DHT and Adiol correlations were nonsignificant. Significant correlations were observed, however, between
all three fecal androgens and bulbourethral gland weight. These
studies demonstrate that fecal T is a valid indicator of reproductive status in the male southern hairy-nosed wombat, with
significant correlations observed between fecal T, plasma T, and
prostate and bulbourethral gland weights. These findings have
important implications for the study of the reproductive endocrinology of the critically endangered northern hairy-nosed
wombat.
The noninvasive method of fecal steroid analysis has
been used successfully to monitor reproduction in a variety
of eutherian mammals [1]. However, fecal steroid analysis
has only been investigated in a few noneutherian species
[2–4]. This is surprising as fecal steroid analysis would
undoubtedly be a valuable research tool in marsupials, especially in those species in which low-stress, noninvasive
sampling methods are desirable. The critically endangered
northern hairy-nosed wombat (Lasiorhinus krefftii) provides a perfect example of one such marsupial [5]. Efforts
to increase the limited knowledge on reproduction in this
species have been hindered by its cryptic nature, difficulties
associated with regular field capture, and the low number
of individuals in the remaining population (approximately
70) [6, 7].
To circumvent the problems of dealing with the endangered northern hairy-nosed wombat population, present
studies have investigated the endocrinology of the more
common, yet closely related, southern hairy-nosed wombat
(Lasiorhinus latifrons). Studies in the southern hairy-nosed
wombat have revealed marked seasonal variations in concentrations of reproductive hormones (androgens) in plasma, as well as the testis and accessory glands in male wombats [8, 9]. Furthermore, the peak reproductive condition of
these male wombats, as determined by plasma androgen
concentration and accessory gland sizes (bulbourethral
gland, prostate weight), coincided with the times of seasonal reproductive activity in female wombats ([9]; unpublished observations).
In this study we investigated whether seasonal differences in plasma androgens (testosterone, dihydrotestosterone, 5a-androstane-3a,17b-diol) could be detected in the
fecal pellets of male southern hairy-nosed wombats. The
objective was to determine whether reproductive status in
the southern hairy-nosed wombat could be determined by
fecal steroid analysis. This study demonstrates that androgens can be measured in fecal pellets, and furthermore that
concentrations of fecal androgens, particularly testosterone,
are indicative of male reproductive status.
male reproductive tract, male sexual function, prostate, seasonal
reproduction, steroid hormones, testosterone
MATERIALS AND METHODS
The financial assistance of the Endangered Species Unit, Environment
Australia, and the Queensland Department of Environment and Heritage
is gratefully acknowledged.
2
Correspondence: Peter Temple-Smith, Director, Conservation and Research, Zoological Parks and Gardens Board, P.O. Box 74, Parkville, Victoria 3052, Australia. FAX: 61 3 9285 9346; e-mail: [email protected]
1
Sample Collection
Fresh fecal samples were collected from two captive
adult southern hairy-nosed wombats (L. latifrons) maintained in outdoor sheltered enclosures at Animal Services,
Monash University, Victoria. Animals were fed a diet consisting of wallaby pellets supplemented with lucerne cubes.
Fecal and plasma samples were collected in the field from
wild southern hairy-nosed wombats (L. latifrons) in the
Swan Reach region of South Australia between 1994 and
Received: 18 December 1998.
First decision: 20 January 1999.
Accepted: 30 March 2000.
Q 2000 by the Society for the Study of Reproduction, Inc.
ISSN: 0006-3363. http://www.biolreprod.org
526
ANDROGENS IN HAIRY-NOSED WOMBATS
527
TABLE 1. Recoveries of 3H-androgens (as % of 3H added) after extraction
and HPLC separation of fecal pellets of wild southern hairy-nosed wombats collected during the breeding (Br) (n 5 7) or nonbreeding (NonBr)
n 5 7) season.
3H-T
Br
Mean
SD
Combined mean
SD
3H-DHT
NonBr
31.4
36.8
26.5
50.1
20.0
53.7
10.8
52.8
34.7
39.6
28.0
60.6
28.5
38.5
25.7
47.4*
8.0
9.2
36.6
14.0
Br
NonBr
22.9
23.3
15.6
39.9
16.8
39.7
6.1
40.9
21.5
28.1
17.0
52.9
16.1
29.9
16.6
36.4*
5.4
10.0
26.5
12.9
3H-Adiol
Br
NonBr
20.6
24.7
19.8
37.5
13.1
39.1
8.0
39.4
20.3
27.5
19.0
47.8
17.6
23.0
16.9
34.1*
4.7
9.2
25.5
11.4
* P , 0.05, Student’s t-test.
1997. Fecal samples were removed directly from the rectum
and sigmoid colon of recently killed animals and stored at
2208C. Matching plasma samples were collected immediately after death via cardiac puncture, centrifuged, and
stored in aliquots at 2208C prior to analysis. The prostate
and bulbourethral glands were dissected from each animal,
fixed in Bouins fluid, and then weighed. All samples were
categorized by collection date in terms of breeding (August–November) and nonbreeding season (February–April),
as defined by Gaughwin et al. [9].
All experiments and field studies were conducted in accordance with the ethics committee guidelines of Monash
University.
Extraction of Fecal Androgens
Fecal samples were thawed, sectioned, and dried in an
incubator at 378C for 48 h. Once dried, samples were
weighed and any large pieces of plant material were removed, following which a number (about four) of fecal
pellets from each sample set was mixed and crushed together to account for potential inter- and intrasample variations in androgen levels. To follow steroid recoveries
throughout processing, 5000 cpm radiolabeled [1a,2a-N3H]testosterone (T), [1,2-N-3H]dihydrotestosterone (DHT),
and [9,11-N-3H]5a-androstane-3a,17b-diol (Adiol) (DuPont, New England Nuclear, Sydney, Australia; specific activities 40–60 Ci/mmol) were added to 0.2 g (dry weight)
of sample. Fecal samples were rotary mixed with 4 ml ether
for 10 min and then allowed to stand at room temperature
for 15 min. The ether fraction was then separated and dried
under nitrogen. The remaining pellets were washed with a
further 2 ml ether, and the rotary mix and drying steps were
repeated. The combined ether fractions were resuspended
in 2 ml of 0.1% (v:v) trifluoroacetic acid (TFA), 20% (v:
v) acetonitrile, and loaded onto a Sep-pak C18 disposable
cartridge (Millipore Waters, Milford, MA). The steroids
were eluted with 0.1% (v:v) TFA, 60% (v:v) acetonitrile,
lyophilized, resuspended in 200 ml HPLC buffer (0.1% [v:
v] TFA, 40% [v:v] acetonitrile), and centrifuged (30 min,
1000 3 g, 48C) prior to HPLC separation.
Extraction of Plasma Androgens
Frozen plasma was thawed, and a 1-ml aliquot was vortexed vigorously for 20 sec in 2 ml of 0.1% (v:v) TFA,
60% (v:v) acetonitrile to which 5000 cpm of radiolabeled
[1a,2a-N-3H]T, [1,2-N-3H]DHT, and [9,11-N-3H]Adiol had
FIG. 1. Correlations between androgen concentrations (testosterone [a],
dihydrotestosterone [b], and Adiol [c]) in plasma (ng/ml) and feces (ng/g
dry weight) of wild southern hairy-nosed wombats (n 5 14).
been added prior to mixing. Plasma samples were then centrifuged (20 min, 6750 3 g, 48C), and the resultant supernatant was diluted 1:3 in 0.1% (v:v) TFA. Samples were
then purified on Sep-pak C18 cartridges and prepared for
HPLC separation, as described above for fecal androgen
extraction.
HPLC Separation and RIA Quantification of Fecal
and Plasma Androgens
The HPLC system consisted of a Waters M6000A HPLC
pump linked to a Waters U6K injector, and the system was
528
HAMILTON ET AL.
0.1 M PBS [0.154 M NaCl], pH 7.4). The RIA employed
was as previously described [10], using a sheep antitestosterone primary antibody (Cox 0457, Sirosera, Sydney, Australia) diluted 1:400 000 in 1:800 normal sheep serum. The
cross-reactivity of this antiserum (provided by the supplier)
was T, 100%; DHT, 56.7%; Adiol, 17.0%; and estrogens or
progestins: ,0.25%. The tracer was an in-house iodinated
histamine-testosterone (10 000 cpm/100 ml). T, DHT, and
Adiol (Sigma Chemical Co., St. Louis, MO) were dissolved
in 100% ethanol and then diluted in assay buffer and used
as standards. The between-assay variations for T, DHT, and
Adiol were 4%, 11%, and 14%, respectively; and the within-assay variations were 10%, 10%, and 13%, respectively.
Typical standard curves ranged from 2.3–152 pg/100 ml for
T (ED50 18 pg/100 ml), 3–365 pg/100 ml for DHT (ED50
28 pg/100 ml), and 14–3000 pg/100 ml for Adiol (ED50 100
pg/100 ml).
Data Analysis
Least-square regression was used to determine levels of
correlation. Seasonal data are represented as mean and standard deviation of the mean. For the determination of statistical significance, a single factor analysis of variance was
conducted. Comparisons of data between groups were conducted using a two-sample Student’s t-test with F-test for
variance.
RESULTS
FIG. 2. Comparisons between breeding (Br) and nonbreeding (non-Br)
seasons for levels of fecal and plasma testosterone (a), dihydrotestosterone
(b), and Adiol (c). Data are expressed as mean 6 SD (n 5 7 per season).
*P , 0.05, ANOVA.
controlled by a Waters Automated Gradient Controller. A
mBondapak C18 column (30 cm 3 0.39 cm; Waters) plus
C18 guard column (Activon, Pennant Hills, Australia) were
equilibrated in HPLC buffer for 30 min at a flow rate at 1
ml/min.
Samples were loaded onto the column using a 250-ml
glass syringe, and fractions (0.5 ml) were collected with a
Pharmacia (Uppsala, Sweden) Frac-100 fraction collector.
To determine 3H steroid recoveries following separation, 50
ml of each fraction was added to 2 ml scintillation fluid
(Packard Emulsifier Safe, Meriden, CA) and counted for 10
min/vial in a beta counter (model 2500TR; Packard). Fractions chromatographing with similar retention times as T,
DHT, and Adiol were then pooled (final volume 1.5–2 ml)
and dried down overnight.
Prior to the measurement of androgens by RIA, samples
were dissolved in 1 ml assay buffer (0.1% [w:v] gelatin in
A baseline–baseline separation of the three tritiated androgens (T, DHT, and Adiol) was achieved by HPLC fractionation (data not shown), with the retention times for each
steroid being 15 min for T, 24.5 min for Adiol, and 27 min
for DHT. The yields of radioactive steroids added as internal standards to fecal pellets collected in the wild from
male southern hairy-nosed wombats and extracted and
chromatographed as described in Materials and Methods
were T 36.6 6 14.0%, DHT 26.5% 6 12.9%, and Adiol
25.5% 6 11.4% (mean 6 SD, n 5 14) (Table 1). Similarly,
the yields of 3H-androgens from plasma samples were T
57.6 6 6.0%, DHT 52.5 6 8.0%, and Adiol 50.8 6 7.9%
(mean 6 SD, n 5 14) (data not shown). Androgen concentrations in plasma and fecal samples as determined by RIA
(see below) were therefore corrected for these losses. For
the purposes of analysis, fecal pellets were then classified
into either breeding season (collected between August–November) or nonbreeding season (collected between February–April), based on the criteria defined by Gaughwin et
al. [9]. When the recoveries of 3H-androgens from fecal
pellets were grouped in this manner, significant (P , 0.05)
differences in recovery between seasons were observed
(Table 1) for all three androgens. No significant differences
were observed for the recoveries of 3H-androgens from the
plasma of animals from the two seasonal groups.
Preliminary experiments were carried out in order to determine the effects of drying temperature on the yields of
radioactive androgens. There were no significant differences in the recoveries of any of the androgens when 3H-labeled steroids were added to fecal pellets prior to the drying
process at either 258C, 378C, or 608C for 48 h (data not
shown).
The concentrations of androgens in fecal pellets from 14
wild southern hairy-nosed wombats as determined by RIA
varied within the range 6.6–25.0 ng/g dry weight for T, 4.0–
24.2 ng/g dry weight for DHT, and 0–34.8 ng/g dry weight
for Adiol. For each androgen, a highly significant (T, P ,
529
ANDROGENS IN HAIRY-NOSED WOMBATS
TABLE 2. Fecal T levels, prostate, and body weights for wombats collected during the breeding and nonbreeding seasons.
Animal number
155
164
175
179
186
191
195
Mean
SD
256
271
278
281
283
537
539
Mean
SD§
Date collected*
Season†
1/9/94
2/9/94
1/10/94
1/10/94
2/10/94
2/10/94
2/10/94
B
B
B
B
B
B
B
23/2/95
21/4/95
22/4/95
22/4/95
22/4/95
7/2/97
7/2/97
NB
NB
NB
NB
NB
NB
NB
Fecal T
(ng/g dry weight)
Body weight
(kg)
Bulbourethral gland
(g)‡
Prostate gland
(g)‡
24.9
13.8
19.5
15.6
14.0
25.0
12.8
17.9
5.28
13.7
7.55
7.18
8.63
6.57
14.0
11.6
9.89
3.15
27.0
30.2
27.2
28.5
29.5
28.2
29.3
28.6
1.19
26.6
25.8
22.7
25.6
27.4
32.2
27.5
26.8
2.87
63.3
51.2
28.8
26.3
38.6
57.7
31.3
42.5a
14.9
13.3
16.0
5.9
15.7
32.7
37.3
32.5
21.9b
12.0
29.6
17.6
21.1
18.5
24.5
31.0
24.3
23.8c
5.18
7.58
10.3
2.95
12.1
15.5
21.1
13.9
11.9d
5.83
* Day/month/year.
† B 5 breeding; NB 5 nonbreeding.
‡ a versus b, c versus d, P , 0.05.
§ Upper limit on 95% confidence interval (P 5 0.05) for fecal T in nonbreeding animals 5 17.6 ng/ml.
0.002; DHT and Adiol, P , 0.001) linear correlation was
observed between plasma and fecal androgen concentrations (Fig. 1). When individuals were grouped by season,
significant (P , 0.05) differences between breeding and
nonbreeding season were observed for both plasma and fecal T (Fig. 2a), whereas no significant differences were seen
for fecal and plasma DHT or Adiol (Fig. 2, b and c), due
to large sample variation within these groups. The upper
limit on the 95% confidence level (P 5 0.05) for fecal T
in nonbreeding animals was 17.6 ng/ml (Table 2), and
based on this parameter three of the seven animals in the
breeding group (numbers 155, 175, and 191) yielded fecal
T levels that were significantly greater than the nonbreeding
group. For all androgens, the mean fecal and plasma androgen concentrations were higher during the breeding season than the nonbreeding season (Fig. 2 and Table 2).
In terms of the effects of androgens on the male accessory glands, a highly significant (P , 0.001) correlation
was observed between fecal T concentration and prostate
weight (Fig. 3a); however, no correlations were seen between the levels of fecal DHT (Fig. 3b) or fecal Adiol (Fig.
3c) and prostate weight. Similarly, prostate weight correlated significantly (P , 0.01) with plasma T, whereas no
correlations were observed for plasma DHT or Adiol and
prostate weight (data not shown). Significant correlations
were observed between the concentrations of all three fecal
androgens and bulbourethral gland weight (T, r 5 0.75, P
, 0.001; DHT, r 5 0.64, P , 0.01; Adiol, r 5 0.50, P ,
0.05; n 5 14). A comparison of breeding and nonbreeding
season weights for the prostate and bulbourethral glands
showed significant (P , 0.05) differences in these parameters (Table 2). When examined in terms of seasonality (Table 2), the data in Figure 3a also showed a clear relationship
between prostate weight, fecal T, and breeding season.
DISCUSSION
Problems such as the cryptic nature, difficulties of field
capture for repeated observations and sampling, and low
remaining population numbers of Australia’s rarest marsupial, the northern hairy-nosed wombat (L. krefftii), have
hindered the acquisition of knowledge on reproduction in
this species. The development and application of a noninvasive sampling technique such as fecal steroid analysis to
monitor reproductive status in wombats is of considerable
importance to future conservation management strategies
for this species. The results from this study show that androgens can be successfully measured in extracts of fecal
pellets collected from the related southern hairy-nosed
wombat, and furthermore that fecal androgens, particularly
testosterone, are indicative of reproductive status in male
wombats as shown by significant correlations with androgen-dependent accessory glands such as the prostate and
bulbourethral glands.
Previous studies in southern hairy-nosed wombats have
found that the peak number of births in this species coincided with seasonal changes in total plasma androgen concentrations (which includes T and DHT) in males and in
the size of the prostate and bulbourethral glands [8, 9].
Similar relationships have been shown in other seasonal
breeding marsupials. For example, several species of marsupial mice, Antechinus stuartii and Antechinus minimus,
have low to undetectable plasma androgen levels and undeveloped prostates during the prebreeding season, followed by significant increases in plasma androgen prior to
and during the breeding season and associated increases in
prostate weight [11–13]. Significant relationships have also
been documented between seasonal changes in peripheral
testosterone concentrations and prostate size in the brushtail
possum [14] and tammar wallaby [15] and bulbourethral
gland size in the tammar wallaby [15]. Interestingly, no
significant changes in testicular weights between seasons
were observed in these species [15, 16]. These reports are
consistent with the findings of the present study in wombats
that has demonstrated that significant differences were
found in concentrations of fecal and plasma androgens between seasons, with a strong correlation between fecal testosterone and prostate or bulbourethral gland weights.
Whether testicular weights in wombats would also show a
similar correlation with plasma or fecal testosterone across
the breeding/nonbreeding season remains unknown; however, recent data [9] have demonstrated a reduction in testicular size and spermatogenic potential of the southern
hairy-nosed wombat in the nonbreeding season.
530
HAMILTON ET AL.
FIG. 3. Correlations between prostate weight and androgen concentrations (testosterone [a], dihydrotestosterone [b], and Adiol [c]) in fecal pellets (ng/g dry weight) of wild southern hairy-nosed wombats (n 5 14).
Fecal androgens were from pellets collected either during the breeding
(solid squares) or nonbreeding seasons (open squares).
The effect of seasonality on fecal androgens was examined in this study by applying the criteria defined by
Gaughwin et al. [9] for wombat reproductive activity to the
selection of field samples. Hence, tissue or serum/fecal
samples were collected over two restricted time intervals in
September/October (breeding) or February/April (nonbreeding). However, based on the hormonal data from the
current study, it is clear that the potential exists to define
seasonality/reproductive activity in the male southern
hairy-nosed wombat in terms of fecal testosterone concen-
trations. This parameter would be a more direct indicator
of wombat reproductive activity by acccounting for the impact of environmental stresses, such as drought, that may
affect the onset of reproduction but that are not accounted
for by a simple calendar-based criterion. The upper limit to
the 95% confidence interval (P 5 0.05) for fecal T in the
nonbreeding wombats was 17.6 ng/ml, and three animals
in the breeding population were indeed above this value,
but the remaining four animals were not. Contributing factors to this lack of discrimination could be the small group
sizes (n 5 7) used in this study, and/or a heterogeneity in
animal response, particularly in the nonbreeding group. For
example, in the nonbreeding group that spans the period
February to April, one male (#537) collected early in February had a breeding size prostate and a fecal T level at
the lower end of the breeding range, while another February
male (#256) had a fecal T level just above the lower limit
of the 95% confidence interval for the breeding group but
a small prostate. The third February male (#539) had a
small prostate and a fecal T level that was below the 95%
confidence interval for breeding males. This suggests that
the transition from breeding to nonbreeding condition for
male southern hairy-nosed wombats at the Swan Reach
field site occurs during the February period. Other studies
have demonstrated that the peak reproductive state of male
southern hairy-nosed wombats coincides with the timing of
ovulation in female wombats, suggesting that either the reproductive activity of the male is determined by that of the
female, or that the same environmental cue(s) influence
both sexes [9]. Whichever is true, the intriguing possibility
occurs that measurement of fecal androgen concentrations
in males will also provide a useful indication of the reproductive status of female wombats.
The significant correlation between fecal testosterone
and prostate weight suggests that, of the three androgens
selected, fecal testosterone provides the best indicator of
reproductive status in male southern hairy-nosed wombats.
In contrast, all fecal androgens showed a strong positive
correlation with bulbourethral gland weight in breeding
males. This suggests that the bulbourethral glands may be
a better indicator of male reproductive status in this species
than the prostate. This conclusion is supported by field observations that show a visible, and perhaps measurable, increase in the prominence of the pericloacal region overlying
the bulbourethral glands in breeding season males (unpublished data). The lack of correlation between fecal DHT
and prostate weight may be related to the limited number
of animals in this study but is consistent with other studies
in humans [17, 18] that show that blood DHT concentrations are not a reliable indicator of peripheral DHT formation. Further, most peripheral DHT is metabolized locally before entering the circulation [18].
The yields of radioactive androgens added to fecal pellets compared with plasma samples were significantly decreased following extraction and HPLC separation, and a
significant yield difference was also observed for fecal pellets from breeding and nonbreeding seasons. This may be
explained by the differences in texture and composition in
feces collected from wild wombats between seasons and is
likely to be the result of seasonal changes in diet [19]. Fecal
samples collected during the nonbreeding season (February–April) were easily crushed to a fine powder prior to
solvent extraction, resulting in an easier separation of the
ether fraction containing the steroids at that stage. However,
a large plant/fibrous component was evident in fecal samples collected from wild wombats during the breeding sea-
ANDROGENS IN HAIRY-NOSED WOMBATS
son, making the ether separation more difficult and resulting in lower recoveries. Previous studies in eutherian mammals have demonstrated, however, that the relationship between fecal and serum hormone profiles is relatively
independent of dietary fiber [1, 20]. Future studies on captive wombats maintained on identical or different diets during the breeding and nonbreeding seasons may provide a
way of examining the effects of diet on the extraction and
yield of fecal androgens in this species.
With further development, fecal steroid analysis has immense potential in the remote monitoring of the reproductive status of hairy-nosed wombats, in particular the northern hairy-nosed wombat, and other marsupials in the wild
and captivity. This development process should include an
assessment of the potential for environmental microbial metabolism or breakdown of androgens from fecal pellets in
the field, as the fecal samples used in this study were collected directly from the rectum and sigmoid colon. Of major interest would be the validation of an analagous technique for the quantitation of female sex steroids, such as
progesterones as successfully applied to other species [1,
3], for use in hairy-nosed wombats. This would provide
important information on the reproductive dynamics of the
female population, including whether individuals were cycling. Furthermore, fecal steroid analysis of wombat feces
could potentially be combined with individual identification
using fecal DNA typing to provide invaluable information
on the reproductive activity and cycles of individual field
animals without the need for capture. The development and
application of this combination of approaches may provide
a way of circumventing the current difficulties faced in determining the reproductive status and activity of northern
hairy-nosed wombats.
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
ACKNOWLEDGMENTS
14.
We thank Ron, Jason, and Ingrid Dibbens for their invaluable assistance in providing support for the field work; John and Joan McCaughley
for access to the wombat population on their property at Swan Reach in
South Australia, for their continuing interest in the project, and for use of
their facilities; and the South Australian Department of Environment, Heritage and Aboriginal Affairs for providing permits to study southern hairynosed wombats in South Australia. All samples were collected on culling
permits issued to John McCaughley by the South Australian Department
of Environment, Heritage and Aboriginal Affairs. The authors also gratefully acknowledge assistance with this study from Assoc. Prof. David Robertson (Prince Henry’s Institute of Medical Research), and technical discussions about steroid extraction with Assoc. Prof. Grant Stone, Department of Animal Science, University of Sydney, NSW, Australia.
15.
16.
17.
18.
19.
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